The host specificity of enterotoxigenic Escherichia coli (ETEC) strains is conferred by the attachment mechanism of these bacteria to the small bowel mucosa. This attachment has been shown to be pili-mediated in strains causing diarrheal disease in animals where binding is associated with the K88 and K99 antigens. The study of the attachment of human-specific ETEC requires human small intestinal cells as an appropriate model system. We have obtained small intestinal cells from healthy, long-term ileostomy patients, by lavage and scraping of the ileal mucosa. Initial results using a radioactively labeled bacteria in a binding assay indicate that human ETEC strains attach to these human intestinal cells. Following the establishment of optimal binding conditions, a wide variety of organisms including human ETEC, animal ETEC, and nontoxigenic E. coli strains will be tested in this adherence model. Several types of pili have been implicated in bacterial attachment to eukaryotic cells: type 1 pili, CFA pili, and the attachment pili isolated from human ETEC in our laboratory. Isolated and purified pili of these recognized types will be used to measure binding to human intestinal cells, to determine the relationship of these pili types and colonization of the human small intestine. We will also continue to develop antisera against the attachment pili isolated from ETEC. We currently find 3 serologically distinct pili types on human ETEC strains. Antisera developed against these 3 types react with 56% of the human ETEC strains in our collection, but no fecal E. coli from healthy adults and animal ETEC strains. The overall goal of these studies is to define the mechanism of attachment of ETEC to human intestinal mucosal cells.